DNA (deoxyribonucleic acid) sequencing has been known for many years. The basic concept of identifying the building blocks of the nucleic acid, containing genetic instructions used in the development and functioning of all known living organisms, has been extended from the discovery of codes to the desire to use genetic information to tackle disease.
The main role of DNA molecules is the long term storage of information. Among other functions, it contains instructions needed for construction of components of cells, in segments referred to as genes. Chemically, DNA consists of two long polymers of simple units called nucleotides, the two strands running in opposite directions to each other. Backbones between the two strands are made of sugars and phosphate groups joined by esther bonds. Attached to each sugar is one of four types of molecules called bases, of type A, C, G or T. It is the sequence of these four bases along the backbone which encodes information. By identification of these bases and their sequence, much information can be derived.
The Sanger method has been, for many years, the standard DNA sequencing technique. This method is very expensive and time consuming. Further techniques have been developed in an effort to make sequencing more efficient and affordable.
Many of the new techniques rely on fluorescent imaging for identification of the bases, known as base calling. A fluorescent moiety is attached to one particular kind of base. The fluorescence in the nucleotide is effected by absorption of light at known wavelength. The fluorescence occurs at another, slightly different, known wavelength. Detection of the fluoresced light indicates the presence of a particular base. Single color fluorescent systems exist wherein different fluids comprising sequencing reagents are washed over a sample in succession and fluorescence indicates the presence of different DNA bases in the DNA sample. Another fluorescent imaging technique is known as four color fluorescence as four different wavelengths of light are used, thereby permitting four types of nucleotide (needed for sequencing reactions) to be present in the sequencing device at the same time. Thus fluid exchanges in the device (which are very slow) can be reduced or kept to a minimum.
A type of four color fluorescent imaging is described in U.S. patent application US 2006/0139634. Other work by Lewis et al (PNAS, 2005, Vol 102, No 15, p 5346) details a temporal color flashing technique which uses continuous wave lasers, blocking filters and complex alignment optics to achieve detection of a given DNA base. Further effort has concentrated on fluorescent decay time constant discrimination, such as Zhu et al (Anal. Chem., 2003, Vol 75, No 10, p 2280). Further progress in gene sequencing is desired as the sequencing of a whole genome or re-sequencing for diseases such as cancer would have huge medical benefits.
A problem with the devices and methods described above is the slow throughput time for obtaining the sequencing information.